The production of hemagglutinins is a well established virulence factor for a number of bacterial species. These include some of the more virulent and troublesome microorganisms that afflict the human host such as Vibrio cholerae, Salmonella, Bordetella pertussis and Escherichia coli. It is thus reasonable to expect that the hemagglutinin of Porphyromonas gingivalis is also involved in virulence. Although it was originally assumed that P. gingivalis has only one hemagglutinin, during the previous years of this grant we have cloned five genes which encode hemagglutinins. The major goal of the experiments proposed in this application is to determine the importance and function of each of the five hemagglutinin genes to the virulence of P. gingivalis. In order to establish the importance of any or all of these genes to virulence, we propose to test isogenic strains mutated in one or more hag genes for virulence in animal model systems. In order to better understand their function, we propose to more completely characterize the genes and their products and to study the regulation of expression of the genes. Thus the hypothesis to be tested by the experiments proposed in this application is: as with genes encoding hemagglutinins of other pathogenic bacterial species, the hag genes of P. gingivalis encode proteins that are involved in virulence/disease activities of this species. The specific aims thus include the following: 1) To construct isogenic mutants of each of the hag genes and test the mutants for virulence-associated activities in appropriate animal and in vitro models. These models include murine abscess and bone loss animal models and in vitro assays of adherence. 2) To assay and quantitate the expression of each of the hag genes in response to various in vitro environmental conditions using fusion/reporter gene constructs. 3) To characterize the hemagglutinin proteins including a) the determination of the variability and significance of the repeat sequence of hagA and b) the mapping and identification of functional domains of the hemagglutinins.